This proposal, entitled, Molecular Mechanisms of Leptin Receptor/Jak2 Action, is an application for competitive renewal of DK56731. The long-term outlook of our previous and future studies is to understand mechanisms of LepRb signaling and to determine how LepRb signals contribute to the regulation of neural function and thence to the control of energy balance, glucose homeostasis, and neuroendocrine function. LepRb mediates tyrosine phosphorylation (Tyr(P))-dependent signaling by means of an associated Jak2 tyrosine kinase. Leptin binding stimulates the Tyr(P) of Jak2 and tyrosine residues on LepRb; each Tyr(P) site mediates a unique complement of intracellular signals. To this point, we have defined the mechanisms of LRb/Jak2 interaction, defined the function of Jak2 Tyr(P) sites, and examined the biology of LepRb Tyr1138AESTAT3 signaling and LepRb Tyr985AESHP2/SOCS3 signaling. Recently, we defined a third Tyr(P) site on LepRb (Tyr1077), which regulates STAT5 signaling and potentially other LepRb signals, and have generated novel mouse models to probe the function of Jak2 and Tyr1077 in LepRb action in vivo. Preliminary data suggest that contributions from LepRb phosphorylation sites are required for most known leptin effects, including some actions that are independent of Tyr1138 and Tyr985. In contrast, Tyr1077 is crucial to the regulation of glycemic control and potentially other physiologic leptin effects. In the context of a variety of data that suggest the importance of the acute (non-transcriptional) effects of leptin in the short-term regulation of energy balance and glycemic control, these data suggest the hypothesis that LepRb Tyr1077 mediates cellular signals required for the acute effects of leptin. In addition to testing this core hypothesis, the proposed research will define the signals and mechanisms by which leptin mediates a variety of physiologic effects and by which leptin modulates blood glucose levels. We propose to: (1) Understand the roles for LepRb signals in the regulation of physiology, focusing on Jak2 and Tyr1077. (2) Define the leptin-mediated regulation of neural function in mouse models of altered LepRb signaling. (3) Determine the signaling mechanisms by which LepRb Tyr1077 and/or other Jak2 and LepRb Tyr(P) sites control physiology and neural function. This approach will reveal the molecular underpinnings of leptin action and delineate the role of each leptin signal in the regulation of neural and organismal physiology by leptin. The mechanistic insights derived from these studies will suggest potential molecular targets for therapeutic intervention in metabolic disease. PUBLIC HEALTH RELEVANCE: Leptin is a key regulator of body energy homeostasis and metabolism, and impaired leptin action may contribute to a variety of metabolic diseases. Understanding the molecular mechanisms of leptin action is thus crucial for our understanding of processes that may be dysregulated in metabolic diseases, as well as for defining potential targets for therapeutic intervention. We have thus been working to define the mechanisms by which the leptin receptor, LepRb, mediates cellular signaling and to understand how each of these signals contributes to the physiologic actions of leptin in vivo. While LepRb Tyr1138/STAT3 signaling is crucial for long-term energy balance, this signaling pathway fails to explain many important aspects of leptin action. LepRb Tyr985, which mediates feedback inhibition on LepRb to attenuate leptin action in vivo, cannot account for residual leptin action. Recent data from our ongoing analysis suggests important roles for LepRb Tyr1077 in leptin action in vivo; signals emanating directly from Jak2 may contribute, as well. We will thus analyze the role of Jak2 and LepRb Tyr1077 in the neural and physiologic actions of leptin by utilizing mouse models in which we have altered LepRb to specifically affect those signals. We will furthermore utilize a set of in vivo systems to define the molecular mediators that lie downstream of Jak2 and/or LepRb Tyr1077 in the regulation of these processes. Overall, these studies will define crucial mediators of leptin action and reveal specific mechanisms by which leptin controls particular physiologic endpoints.